1. Field of Invention
This invention relates to a confocal observation system; and more particularly, to an optical image separation system connected to a confocal image output port of a Nipkow disk type confocal scanner.
2. Description of the Prior Art
FIG. 1 shows a conventional Nipkow disk type confocal scanner used with a microscope, wherein confocal scanner 100 is connected to microscope 200. Illuminating parallel excitation light beam 1 (depicted as a chain line) is converged into individual beams by micro lens array disk (called xe2x80x9cML diskxe2x80x9d) 2, and passed through pinholes of pinhole array disk (called xe2x80x9cNipkow diskxe2x80x9d) 4 after being transmitted through dichroic mirror (called xe2x80x9cDMxe2x80x9d) 3. Light beam 1 is then focused onto sample 6 by objective lens 5 of the microscope 200. Fluorescence signal 7 (depicted as a continuous line) emitted from sample 6 is transmitted through objective lens 5 and is focused on the pinholes of Nipkow disk 4. Fluorescence signal 7, passed through the pinholes, is reflected by DM 3 and is caused to form a fluorescence image on a two dimensional sensor 10 by relay lens 9. DM 3 is designed to transmit excitation light 1 and to reflect the desired fluorescence signal 7.
ML disk 2, and Nipkow disk 4 are concurrently turned by rotation shaft 11, both being mechanically connected to each other by member 8. Mico lenses and pinholes formed on Nipkow disk 4 are arranged so that the pinholes scan over plane 12 to observe sample 6. The plane on which pinholes of Nipkow disk 4 are arranged, plane 12 to be observed on sample 6, and light detecting surface of the sensor 10 are arranged in optically conjugate relationship to each other. Accordingly, an optical sectional image of sample 6, that is, a confocal image is formed on sensor 10.
A plurality of specimens placed on the same plane can be simultaneously observed because a Nipkow disk type confocal scanner uses a two dimensional sensor. Also, a confocal image of sample 6 can be formed on the light detecting surface of the sensor 10 in a short time by rotating ML disk 2 and Nipkow disk 4 together at a high speed. This makes it possible to acquire at high speeds confocal images of all of the samples 60, on which many specimens to be inspected are arranged in a matrix as shown in FIG. 2, moving them relative to the microscope and the confocal scanner. Accordingly, confocal scanners are used for screening procedures used in developing new medicine. Furthermore, when a sample emitting sufficiently bright fluorescence signals is used as the object of test or inspection, use of the ML disk 2 is sometimes not necessary.
The conventional confocal scanner has a problem in that the number of samples that can be processed per unit time in the screening procedure of FIG. 2, is limited to those where a sample emits only one type of fluorescence. Disadvantageously, when a multiple staining procedure is used, a plurality of different fluorescence wavelengths, depending on sample reactions to the different wavelengths, are detected. This is done by adding separate fluorescence reagents having different wavelengths of fluorescence for each sample 60, or by adding a plurality of types of fluorescence reagents to all of the samples. In that case, sample 60 having a plurality of different specimen reacting to different fluorescence reagents, is first inspected using a DM 3 that has a specific reflection characteristic and can detect that specific first fluorescence wavelength. Then, after replacing the DM3 with another DM3 having another reflection characteristic corresponding to the second fluorescence wavelength, the entire specimen having the plurality of samples, must again be inspected. As can be appreciated, the conventional confocal scanner can be improved.
As shown in FIG. 3, one way of handling the foregoing problem is to use a plurality of sets of microscopes 200a, 200b, 200c; confocal scanners 100a, 100b, 100c; and two dimensional sensors 10a, 10b, 10c, arranged, respectively, for each fluorescence wavelength. When inspection for fluorescence wavelength xcexn to sample 60n is completed for one wavelength, sample 60n is sent to the next inspections system n+1 comprising microscope 200n+1, scanner 100n+1 and two dimensional sensor 10n+1, and then the process is repeated for the next fluorescence wavelength of the sample. In other words, the inspection for each fluorescence wavelength is repeated using the different sets. However, this arrangement is not satisfactory since it involves use of a plurality of sets of devices and hence cost is expensive, and repeated procedures are time consuming.
Accordingly, an object of the invention is to overcome the aforementioned and other deficiencies, disadvantages and problems of the prior art.
Another object is to provide an optical image separation system connected to a Nipkow disk type confocal scanner, wherein return light from a sample is detected by separating the return light concurrently into a plurality of different wavelength regions without repeating the DM changes, as done in the prior art; and inspection of the total number of specimen by using the entire number of wavelengths, without using a plurality of sets of scanners and microscopes, as done in the prior art.
The foregoing and other objects are attained in the invention which encompasses an optical image separation system connected to a confocal image output port of a Nipkow disk type confocal scanner, wherein the optical image separation system comprises a return light separating means that separates the light returned from a sample and emitted from the output port of the scanner into light beams of a plurality of wavelength regions or a plurality of portions of the same wavelength region.
The various features, aspects, and advantages and effects of the invention are further detailed hereinbelow.